• BMB Reports
• /
• v.34 no.6
• /
• pp.517-525
• /
• 2001

Six renaturable protein kinases that utilize the myelin basic protein (MBP) as a substrate were activated during prolonged exposure of cardiac myocytes to okadaic acid (OA). We characterized the substrate preference and activation of these kinases, with particular emphasis on 3 novel kinases-MBPK-55, MBPK-62 and MBPK-87. The transcription factors c-Jun, Elk, ATF2, and c-Fos that are used to assess mitogen-activated protein kinase activation were all poor substrates for these three kinases. MAPKAPK2 was also not phosphorylated. In contrast, Histone IIIS was phosphorylated by MBPK-55 and MBPK-62. These protein kinases were activated in cultured cardiac fibroblasts, H9c2 cardiac myoblasts, and Cos cells. High concentrations (0.5 to $1\;{\mu}M$) of OA were essential for the activation of the protein kinases in all of the cell types examined, whereas calyculin A [an inhibitor of protein phosphatase 1 (PP1) and PP2A], cyclosporin A (a PP2B inhibitor), and an inactive OA analog all failed to activate these kinases. The high dose of okadaic acid that is required for kinase activation was also required for phosphatase inhibition, as assessed by immunoblotting whole cell lysates with anti-phosphothreonine antibodies. A variety of chemical inhibitors, including PD98059 (MEK-specific), genistein (tyrosine kinase-specific) and Bisindolylmaleimide I (protein kinase C-specific), failed to inhibit the OA activation of these kinases. Thus, MBPK-55 and MBPK-62 are also Histone IIIS kinases that are widely expressed and specifically activated upon exposure to high OA concentrations.

• Journal of Microbiology and Biotechnology
• /
• v.7 no.1
• /
• pp.56-61
• /
• 1997

A strain producing immunosuppressive substances was isolated from a soil in Cheju island. By morphological, cultural, and physiological studies, the strain was identified as Streptomyces lydicus MCY-524. Cultured broth was purified by silica gel, sephadex LH-20 and preparative HPLC and gave two immunosuppressive compounds, MCH-22 and MCH-32. They dramatically suppressed the B cell activation with lipopolysaccharide, T cell activation by mixed lymphocyte response, and primary T-dependent antibody response at a final concentration of 1 ${\mu}g$/ml. They also markedly suppressed the proliferation of lymphocytes induced by lipopolysaccharide, pokeweed mitogen, and concanavaline A at the same concentration. Their suppressive activities, which were comparable to those of cyclosporin A, suggested that they were potent and broad immunotoxic agents on the immune functions of murine lymphocytes.

• Toxicological Research
• /
• v.30 no.3
• /
• pp.193-198
• /
• 2014

Degradation of glucose is aberrantly increased in hyperglycemia, which causes various harmful effects on the liver. Methylglyoxal is produced during glucose degradation and the levels of methylglyoxal are increased in diabetes patients. In this study we investigated whether methylglyoxal induces mitochondrial impairment and apoptosis in HepG2 cells and induces liver toxicity in vivo. Methylglyoxal caused apoptotic cell death in HepG2 cells. Moreover, methylglyoxal significantly promoted the production of reactive oxygen species (ROS) and depleted glutathione (GSH) content. Pretreatment with antioxidants caused a marked decrease in methylglyoxal-induced apoptosis, indicating that oxidant species are involved in the apoptotic process. Methylglyoxal treatment induced mitochondrial permeability transition, which represents mitochondrial impairment. However, pretreatment with cyclosporin A, an inhibitor of the formation of the permeability transition pore, partially inhibited methylglyoxal-induced cell death. Furthermore, acute treatment of mice with methylglyoxal increased the plasma levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), indicating liver toxicity. Collectively, our results showed that methylglyoxal increases cell death and induces liver toxicity, which results from ROS-mediated mitochondrial dysfunction and oxidative stress.

• The Korean Journal of Nuclear Medicine
• /
• v.37 no.6
• /
• pp.382-392
• /
• 2003

Purpose: Uptakes of Tc-99m MIBI (MIBI) and Tc-99m tetrofosmin (tetrofosmin) in human non-small cell lung cancer A549, multidrug-resistance associated protein (MRP) expressing cell, were investigated in vitro and in vivo. Materials and Methods: Western blot analysis and immunohistochemistry were used for detection of MRP in A549 cells with anti-MRPr1 antibody. Cellular uptakes of two tracers were evaluated at $100{\mu}M$ of verapamil (Vrp), $50{\mu}M$ of cyclosporin A (CsA) and $25{\mu}M$ of butoxysulfoximide (BSO) after incubation with MIBI and tetrofosmin for 30 and 50 min at $37^{\circ}C$, using single cell suspensions at $1{\times}10^6cells/ml$. Radioactivities in supernatants and pellets were measured with gamma well counter. A549 cells were inoculated in each flanks of 24 nude mice. Group 1 (Gr1) and Gr3 mice were treated with only MIBI or tetrofosmin, and Gr2 and Gr4 mice were treated with 70mg/kg of CsA i.p. for 1 hour before injection of 370KBq of MIBI or tetrofosmin. Mice were sacrificed at 10, 60 and 240 min. Radioactivities of organs and tumors were expressed as percentage injected dose per gram of tissue (%ID/gm). Results: Western blot analysis of the A549 cells detected expression of MRPr1 (190 kDa) and immunohistochemical staining of tumor tissue for MRPr1 revealed brownish staining in cell membrane but not P-gp. Upon incubating A549 cells for 60 min with MIBI and tetrofosmin, cellular uptake of MIBI was higher than that of tetrofosmin. Coincubation with modulators resulted in an increase in cellular uptakes of MIBI and tetrofosmin. Percentage increase of MIBI was higher than that of tetrofosmin with Vrp by 623% and 427%, CsA by 753% and 629% and BSO by 219% and 140%, respectively. There was no significant difference in tumoral uptakes of MIBI and tetrofosmin between Gr1 and Gr3. Percentage increases in MIBI (114% at 10 min, 257% at 60 min, 396% at 240 min) and tetrofosmin uptake (110% at 10 min, 205% at 60 min, 410% at 240 min) were progressively higher by the time up to 240 min with CsA. Conclusion: These results indicate that MIBI and tetrofosmin are suitable tracers for imaging MRP-mediated drug resistance in A549 tumors. MIBI may be a better tracer than tetrofosmin for evaluating MRP reversal effect of modulators.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.23 no.1
• /
• pp.166-173
• /
• 2009

Cyclosporine(CsA) is an immunosupressant drug widely used in post-allogeneic organ transplant to reduce the activity of the patient's immune system and so the risk of organ rejection. It has been studied in transplants of skin, heart, kidney, liver, lung, pancreas, bone marrow and small intestine. Initially isolated from a Norwegian soil sample, Both kidney and liver dysfunction are prominent side effects of CsA. The present study was designed to determine the possible protective effect of salidroside(Sal) isolated from the BuOH extract of Acer termentosum Max against oxidative damage in CsA-treated(50 mg/kg, ip) nephrotoxicity rats. Results showed oral administration of methanol and butanol extact of Acer termentosum Max(200 mg/kg, po) significantly reduced activities of marker enzymes(BUN, Creatinine) and LDH activity in serum to CsA induced experimental kidney injured rats. And significantly decrcease of protein amount level in urine and activities of free radical formation enzyme were significantly improved by the treatment of Sal. And significantly decrcease of MDA level in kidney and activities of calalase, glutathione peroxidation and SOD were significantly improved by the treatment of Sal(20 mg/kg, po). But glutathione concentration and glutathione S-transferase actitity was not affected. Results of this study revealed that Sal could afford a significant protection in the alleviation of CsA-induced nephrotoxicity injury.

• Molecules and Cells
• /
• v.20 no.1
• /
• pp.142-150
• /
• 2005

Members of the multifunctional Cyp family have been isolated from a wide range of organisms. However, few functional studies have been performed on the role of these proteins as chaperones in red alga. For studying the function of cDNA GjCyp-1 isolated from the red alga (Griffithsia japonica), we expressed and purified a recombinant GjCyp-1 containing a hexahistidine tag at the amino-terminus in Escherichia coli. An expressed fusion protein, $H_6GjCyp-1$ maintained the stability of E. coli proteins up to $50^{\circ}C$. For a functional bioassay for recombinant $H_6GjCyp-1$, the viability of E. coli cells overexpressing $H_6GjCyp-1$ was compared with that of cells not expressing $H_6GjCyp-1$ at $50^{\circ}C$. After high temperature treatment for 1 h, E. coli overexpressing $H_6GjCyp-1$ survived about three times longer than E. coli lacking $H_6GjCyp-1$. Measurement of the light scattering of luciferase (luc) showed that GjCyp-1 prevents the aggregation of luc during mild heat stress and that the thermoprotective activity of GjCyp-1 is blocked by cyclosporin A (CsA), an inhibitor of Cyps. Furthermore, the Cyp-CsA complex inhibited the growth of E. coli under normal conditions. The results of the GjCyp-1 bioassays as well as in vitro studies strongly suggest that Cyp confers thermotolerance to E. coli.

• Journal of Yeungnam Medical Science
• /
• v.26 no.2
• /
• pp.143-147
• /
• 2009

Hemorrhagic cystitis (HC) is a common complication after allogeneic transplantation. Early posttransplant HC occurs in association with cyclophosphamide, while later on HC results from viral infections such as polyomavirus BK (BKV) and adenovirus. We report here the case of a 57-year-old woman who received an instillation of cidofovir into the bladder for the treatment of hemorrhagic cystitis after allogeneic peripheral stem cell transplantation for her acute myeloid leukemia. Cyclophosphamide and busulfan were used as conditioning treatments. Cyclosporin was administered daily. On the 71st day after transplantation, the patient developed acute severe hemorrhagic cystitis, and BK virus was demonstrated in the urine samples using polymerase chain reaction. Her urinary symptoms did not improve in spite of palliative treatment, but a response was evident after intravesical cidofovir treatment.

• Journal of Haehwa Medicine
• /
• v.21 no.1
• /
• pp.11-21
• /
• 2012

New onset diabetes is a major complication after kidney transplantation. However, the natural course of posttransplantation diabetes mellitus (PTDM) remains unclear. The aim of this study was to demonstrate the detailed natural courses of PTDM according to the onset and persistency of hyperglycemia, and to investigate risk factors for development of different courses of PTDM in renal allograft recipients. The purpose of this study is to develop novel immune suppressants for PTDM using of action mechanism of them. The use of immunosuppressive drugs in transplanted patients is associated with the development of diabetes, possibly due to ${\beta}$-cell toxicity. To better understand the mechanisms leading to post-transplant diabetes, we investigated the actions of prolonged exposure of ${\beta}$-cells to therapeutical levels of tacrolimus (FK506) or cyclosporin A(CsA). The immunosuppressive drug cyclosporine(CsA) is a potent agent widely used after organ transplantations and various autoimmune disorders. After using CsA, some patients suffer severe complications including renal and vascular toxicity. The renal or vascular toxicity is influenced by the degree of the endothelial damage. FK506(tacrolimus) is a widely used immunosuppressive agent in the treatment of various medical conditions, including autoimmune disease, bone marrow and organ transplantations. We found some interesting clusters and confirmed the feasibility of cDNA microarray in the study of Immunosuppressant. In this study, we investigated gene expression patterns induced by Immunosuppressant in RIN-m5F of rat insulinoma cell line. Gene expressions evaluated using cDNA microarry in two clusters were increased or decreased. this study provides comprehensive comparison of the patterns of gene expression changes induced by CsA and FK506 in ${\beta}$-cells. This study could establish that the mode of action mechanism by which currently used insulin inhibitors inducing PTDM could be elucidated at least in part, which raises the possibility that novel immune suppressive PTDM can be developed. The molecular biological study on PTDM will also contribute the progress in diabetes research field as well as in that of PTDM.

• Journal of Periodontal and Implant Science
• /
• v.26 no.3
• /
• pp.669-679
• /
• 1996

Gingival overgrowth is a well known side effect of several drugs, including nifedipine, phenytoin, cyclosporin, dilitiazem, verapamil. A number of studies have been performed to investigate the mechanism by which nifedipine(a calcium channel blocking agent) affects the gingival tissue. The aim of the present work was to investigate the effect of nifedipine on healthy gingival fibroblasts with special emphasis on determining the changes in cellular proliferation and protein and collagen synthesis. Gingival fibroblasts were obtained from the explants of healthy gingiva of extracted 3rd molars or premolar teeth extracted from the patients for orthodontic treatment. To evaluate the effect of nifedipine on cell proliferation, the cells were seeded at a cell density of $1{\times}10^4$cells/well in 24-well culture plates and treated with 100 and 200ng/ml of nifedipine for 10days. After trypsinization, the cells were counted with a haemocytometer on 1st, 3rd, 5th, 7th and 10th days. Then, MTT assay was carried out. For total protein and percent collagen synthesis, $3{\mu}Ci/ml$ $^3H-proline$ was added to each well for the final 4 hours of the incubation period. The results indicate that nifedipine does not influence cell proliferation in healthy gingival fibroblast in vitro and has a specific effect in reducing total protein and percent collagen synthesis. On the above the findings, exogenous nifedipine does not influence on healthy human gingival fibroblast proliferation and protein and collagen synthesis.

• Korean Journal of Medicinal Crop Science
• /
• v.19 no.5
• /
• pp.380-387
• /
• 2011

This study was carried out to investigate the inhibitory effect of Angelicae Dahuricae Radix (ADR) extract on chronic obstructive pulmonary disease (COPD) induced by cigarette smoke condensate (CSC) and lipopolysaccharide (LPS) in mice. COPD was induced by intratracheal instillation of LPS and CSC 5 times for 12 days; this increased airway hyperresoponsiveness (AHR) and inflammatory cells in bronchoalveolar lavage fluid (BALF). ADR extract was administered orally at a dose of 50 and 200 mg/kg. The concentration of imperatorin, a major component of ADR and therefore used as a measure of quality control, was $0.098%{\pm}0.018%$. Treatment of the mice with ADR extract (50 and 200 mg/kg) alleviated AHR and reduced inflammatory cell counts. Treatment with cyclosporin A (CSA; 10 mg/kg) also modulated AHR and reduced inflammatory cells effectively. Compared with CSA treatment, treatment with ADR (50 mg/kg) extract reduced neutrophil and $CD4^+/CD3^+$ cell counts by 22.67% and 44.92%, respectively. In addition, compared with CSA treatment, treatment ADR 200 mg/kg reduced neutrophils, $CD4^+/CD3^+$ cells and $CD8^+/CD3^+$ cells, by 32.10%, 83.17% and 82.11%, respectively. These results indicate that ADR extract may have an inhibitory effect on COPD induced by LPS and CSC in mice.